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1.
Angiogenesis ; 17(2): 383-93, 2014 Apr.
Article in English | MEDLINE | ID: mdl-24154862

ABSTRACT

Lymphatic vessels have traditionally been regarded as a rather inert drainage system, which just passively transports fluids, leukocytes and antigen. However, it is becoming increasingly clear that the lymphatic vasculature is highly dynamic and plays a much more active role in inflammatory and immune processes. Tissue inflammation induces a rapid, stimulus-specific upregulation of chemokines and adhesion molecules in lymphatic endothelial cells and a proliferative expansion of the lymphatic network in the inflamed tissue and in draining lymph nodes. Moreover, increasing evidence suggests that inflammation-induced changes in the lymphatic vasculature have a profound impact on the course of inflammatory and immune responses, by modulating fluid drainage, leukocyte migration or the removal of inflammatory mediators from tissues. In this review we will summarize and discuss current knowledge of the inflammatory response of lymphatic endothelium and of inflammation-induced lymphangiogenesis and the current perspective on the overall functional significance of these processes.


Subject(s)
Endothelium, Lymphatic/pathology , Inflammation/pathology , Animals , Endothelium, Lymphatic/metabolism , Humans , Inflammation/immunology , Inflammation Mediators/metabolism , Lymphangiogenesis/genetics
2.
Blood ; 122(13): 2271-81, 2013 Sep 26.
Article in English | MEDLINE | ID: mdl-23963040

ABSTRACT

The cytokine interleukin (IL)-7 exerts essential roles in lymph node (LN) organogenesis and lymphocyte development and homeostasis. Recent studies have identified lymphatic endothelial cells (LECs) as a major source of IL-7 in LNs. Here, we report that LECs not only produce IL-7, but also express the IL-7 receptor chains IL-7Rα and CD132. Stimulation with recombinant IL-7 enhanced LEC in vitro activity and induced lymphangiogenesis in the cornea of wild-type (WT) mice. Whereas in IL-7Rα(-/-) mice, dermal lymphatic vessels (LVs) were abnormally organized and lymphatic drainage was compromised, transgenic overexpression of IL-7 in mice resulted in an expanded dermal LV network with increased drainage function. Moreover, systemic treatment with recombinant IL-7 enhanced lymphatic drainage in the skin of WT mice and of mice devoid of lymphocytes. Experiments in IL-7Rα(-/-) bone marrow chimeras demonstrated that the drainage-enhancing activity of IL-7 was exclusively dependent on IL-7Rα expression in stromal but not in hematopoietic cells. Finally, near-infrared in vivo imaging performed in IL-7Rα(-/-) mice revealed that the pumping activity of collecting vessels was normal but fluid uptake into lymphatic capillaries was defective. Overall, our data point toward an unexpected new role for IL-7 as a potential autocrine mediator of lymphatic drainage.


Subject(s)
Endothelial Cells/metabolism , Interleukin-7/metabolism , Lymphatic Vessels/metabolism , Animals , Flow Cytometry , Fluorescent Antibody Technique , Humans , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout
3.
FASEB J ; 27(3): 978-90, 2013 Mar.
Article in English | MEDLINE | ID: mdl-23169771

ABSTRACT

Adhesion molecules play an important role in vascular biology because they mediate vascular stability, permeability, and leukocyte trafficking to and from tissues. Performing microarray analyses, we have recently identified activated leukocyte cell adhesion molecule (ALCAM) as an inflammation-induced gene in lymphatic endothelial cells (LECs). ALCAM belongs to the immunoglobulin superfamily and engages in homophilic as well as heterophilic interactions. In this study, we found ALCAM to be expressed at the protein level in human and murine lymphatic and blood vascular endothelial cells in vitro and in the vasculature of human and murine tissues in vivo. Functional in vitro experiments revealed that ALCAM mediates adhesive interactions, migration, and tube formation in LECs, suggesting a role for ALCAM in lymphatic vessel (LV) stability and in lymphangiogenesis. Furthermore, ALCAM supported dendritic cell (DC) adhesion to lymphatic endothelium. In agreement with these findings, experiments performed in ALCAM mice detected reduced LEC numbers in various tissues and defects in the formation of an organized LV network. Moreover, DC migration from lung to draining lymph nodes was compromised in ALCAM mice. Collectively, our data reveal a novel role for ALCAM in stabilizing LEC-LEC interactions and in the organization and function of the LV network.


Subject(s)
Antigens, CD/metabolism , Cell Adhesion Molecules, Neuronal/metabolism , Cell Communication/physiology , Cell Movement/physiology , Endothelial Cells/metabolism , Fetal Proteins/metabolism , Lymph Nodes/metabolism , Lymphatic Vessels/metabolism , Animals , Antigens, CD/genetics , Cell Adhesion/physiology , Cell Adhesion Molecules, Neuronal/genetics , Cells, Cultured , Dendritic Cells/cytology , Dendritic Cells/metabolism , Endothelial Cells/cytology , Fetal Proteins/genetics , Humans , Lung/cytology , Lung/metabolism , Lymph Nodes/cytology , Lymphatic Vessels/cytology , Mice , Mice, Knockout
4.
Blood ; 120(24): 4675-83, 2012 Dec 06.
Article in English | MEDLINE | ID: mdl-22955921

ABSTRACT

Nonhematopoietic stromal cells of secondary lymphoid organs form important scaffold and fluid transport structures, such as lymph node (LN) trabeculae, lymph vessels, and conduits. Furthermore, through the production of chemokines and cytokines, these cells generate a particular microenvironment that determines lymphocyte positioning and supports lymphocyte homeostasis. IL-7 is an important stromal cell-derived cytokine that has been considered to be derived mainly from T-cell zone fibroblastic reticular cells. We show here that lymphatic endothelial cells (LECs) are a prominent source of IL-7 both in human and murine LNs. Using bacterial artificial chromosome transgenic IL-7-Cre mice, we found that fibroblastic reticular cells and LECs strongly up-regulated IL-7 expression during LN remodeling after viral infection and LN reconstruction after avascular transplantation. Furthermore, IL-7-producing stromal cells contributed to de novo formation of LyveI-positive lymphatic structures connecting reconstructed LNs with the surrounding tissue. Importantly, diphtheria toxin-mediated depletion of IL-7-producing stromal cells completely abolished LN reconstruction. Taken together, this study identifies LN LECs as a major source of IL-7 and shows that IL-7-producing stromal cells are critical for reconstruction and remodeling of the distinct LN microenvironment.


Subject(s)
Endothelial Cells/metabolism , Interleukin-7/metabolism , Lymph Nodes/metabolism , Stromal Cells/metabolism , Adult , Animals , Cell Line , Cell Proliferation , Cells, Cultured , Female , Fibroblasts/cytology , Fibroblasts/metabolism , Gene Expression , Humans , Immunohistochemistry , Interleukin-7/genetics , Kidney/metabolism , Luminescent Proteins/genetics , Luminescent Proteins/metabolism , Lymph Nodes/embryology , Lymph Nodes/transplantation , Lymphatic Vessels/cytology , Lymphatic Vessels/metabolism , Lymphocytic Choriomeningitis/genetics , Lymphocytic Choriomeningitis/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Microscopy, Confocal , Reverse Transcriptase Polymerase Chain Reaction
5.
Biol Chem ; 392(11): 983-93, 2011 Nov.
Article in English | MEDLINE | ID: mdl-21871011

ABSTRACT

Cathepsin B has been shown to not only reside within endo-lysosomes of intestinal epithelial cells, but it was also secreted into the extracellular space of intestinal mucosa in physiological and pathological conditions. In an effort to further investigate the function of this protease in the intestine, we generated a transgenic mouse model that would enable us to visualize the localization of cathepsin B in vivo. Previously we showed that the A33-antigen promoter could be successfully used in vitro in order to express cathepsin B-green fluorescent protein chimeras in cells that co-expressed the intestine-specific transcription factor Cdx1. In this study an analog approach was used to express chimeric cathepsin B specifically in the intestine of transgenic animals. No overt phenotype was observed for the transgenic mice that reproduced normally. Biochemical and morphological studies confirmed that the overall intestinal phenotype including the structure and polarity of this tissue as well as cell numbers and differentiation states were not altered in the A33-CathB-EGFP mice when compared to wild type animals. However, transgenic expression of chimeric cathepsin B could not be visualized because it was not translated in situ although the transgene was maintained over several generations.


Subject(s)
Cathepsin B/genetics , Intestinal Mucosa/metabolism , Transgenes , Amino Acid Sequence , Animals , Cathepsin B/analysis , Gene Expression , Green Fluorescent Proteins/genetics , Intestines/ultrastructure , Membrane Glycoproteins/genetics , Mice , Mice, Inbred C57BL , Mice, Transgenic , Molecular Sequence Data , Promoter Regions, Genetic
6.
Blood ; 118(1): 205-15, 2011 Jul 07.
Article in English | MEDLINE | ID: mdl-21596851

ABSTRACT

Chemokines and adhesion molecules up-regulated in lymphatic endothelial cells (LECs) during tissue inflammation are thought to enhance dendritic cell (DC) migration to draining lymph nodes, but the in vivo control of this process is not well understood. We performed a transcriptional profiling analysis of LECs isolated from murine skin and found that inflammation induced by a contact hypersensitivity (CHS) response up-regulated the adhesion molecules ICAM-1 and VCAM-1 and inflammatory chemokines. Importantly, the lymphatic markers Prox-1, VEGFR3, and LYVE-1 were significantly down-regulated during CHS. By contrast, skin inflammation induced by complete Freund adjuvant induced a different pattern of chemokine and lymphatic marker gene expression and almost no ICAM-1 up-regulation in LECs. Fluorescein isothiocyanate painting experiments revealed that DC migration to draining lymph nodes was more strongly increased in complete Freund adjuvant-induced than in CHS-induced inflammation. Surprisingly, DC migration did not correlate with the induction of CCL21 and ICAM-1 protein in LECs. Although the requirement for CCR7 signaling became further pronounced during inflammation, CCR7-independent signals had an additional, albeit moderate, impact on enhancing DC migration. Collectively, these findings indicate that DC migration in response to inflammation is stimulus-specific, mainly CCR7-dependent, and overall only moderately enhanced by LEC-induced genes other than CCL21.


Subject(s)
Cell Movement/immunology , Dendritic Cells/immunology , Dermatitis, Contact/immunology , Endothelial Cells/immunology , Lymph Nodes/immunology , Animals , Chemokine CCL21/genetics , Chemokine CCL21/immunology , Chemokine CCL21/metabolism , Dendritic Cells/cytology , Ear, External/immunology , Female , Gene Expression/immunology , Gene Expression Profiling , Intercellular Adhesion Molecule-1/genetics , Intercellular Adhesion Molecule-1/immunology , Intercellular Adhesion Molecule-1/metabolism , Lymph Nodes/cytology , Mice , Mice, Inbred C57BL , Mice, Mutant Strains , Oligonucleotide Array Sequence Analysis , Receptors, CCR7/genetics , Receptors, CCR7/immunology , Receptors, CCR7/metabolism , Up-Regulation/immunology , Vascular Cell Adhesion Molecule-1/genetics , Vascular Cell Adhesion Molecule-1/immunology , Vascular Cell Adhesion Molecule-1/metabolism
7.
Exp Cell Res ; 316(17): 2982-92, 2010 Oct 15.
Article in English | MEDLINE | ID: mdl-20599951

ABSTRACT

The lymphatic vascular system plays an important role in inflammation and cancer progression, although the molecular mechanisms involved are poorly understood. As determined by comparative transcriptional profiling studies of ex vivo isolated mouse intestinal lymphatic endothelial cells versus blood vascular endothelial cells, thymus cell antigen 1 (Thy1, CD90) was expressed at much higher levels in lymphatic endothelial cells than in blood vascular endothelial cells. These findings were confirmed by quantitative PCR, and at the protein level by FACS and immunofluorescence analyses. Thy1 was also strongly expressed by tumor-associated lymphatic vessels, as evaluated in a B16 melanoma footpad model in mice. Blockade of Thy1 inhibited tumor cell adhesion to cultured mouse lymphatic endothelial cells. Importantly, treatment of human dermal microvascular endothelial cells with tumor necrosis factor or phorbol 12-myristate 13-acetate resulted in Thy1 upregulation in podoplanin-expressing lymphatic endothelial cells, but not in podoplanin-negative blood vascular endothelial cells. Moreover, adhesion of human polymorphonuclear and mononuclear leukocytes to human lymphatic endothelial cells was Thy1-dependent. Together, these results identify Thy1 as a novel lymphatic vessel expressed gene and suggest its potential role in the cell adhesion processes required for tumor progression and inflammation.


Subject(s)
Cell Adhesion , Endothelium, Lymphatic/physiology , Lymphatic Vessels/metabolism , Thy-1 Antigens/biosynthesis , Animals , Endothelial Cells/metabolism , Endothelium, Vascular , Humans , Inflammation/pathology , Melanoma, Experimental/pathology , Mice
8.
Biol Chem ; 389(8): 1085-96, 2008 Aug.
Article in English | MEDLINE | ID: mdl-18979632

ABSTRACT

We hypothesized that tissue-specific expression of cathepsin B-enhanced green fluorescent protein (CB-EGFP) can be driven by the A33-antigen promoter that contains positive cis-regulatory elements, including caudal-related homeobox (CDX) binding sites. The intestine-specific transcription factor Cdx1 is crucial for A33-antigen promoter activation and could thereby induce expression of CB-EGFP. This concept was tested by construction of the vector pA33-CathB-EGFP encoding CB-EGFP downstream of the A33-antigen promoter. Its Cdx1 dependence, as an indication of its intestine-specific expression, was tested in Cdx1-negative CHO-K1 cells. Cdx1 expression was achieved upon transfection with pCdx1-DsRed-Express and was indicated by red fluorescence of the simultaneously translated reporter protein. Immunolabeling with Cdx1-specific antibodies showed correct targeting of the transcription factor to its point of action in nuclei of transfected cells. Co-transfection experiments with plasmids pA33-CathB-EGFP and pCdx1-DsRed-Express confirmed the hypothesis that Cdx1 indeed activates CB-EGFP expression in a manner dependent on the A33-antigen promoter. Co-localization with compartment-specific markers and subcellular fractionation confirmed CB-EGFP trafficking along the expected route to endolysosomal compartments. Hence, the A33-antigen promoter represents a potent tool for induction of Cdx1-dependent CB-EGFP expression in vitro. Our proof-of-principle studies confirm the suitability of this approach in visualizing protease transport in Cdx1-positive tissues of the gastrointestinal tract.


Subject(s)
Cathepsin B/metabolism , Gene Expression , Genes, Reporter/genetics , Intestines/enzymology , Animals , CHO Cells , Cathepsin B/genetics , Cricetinae , Cricetulus , Genetic Vectors/genetics , Homeodomain Proteins/genetics , Homeodomain Proteins/metabolism , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Mice , Organ Specificity , Promoter Regions, Genetic/genetics , Protein Transport
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